EP2685232B1 - Method for using object-capturing device - Google Patents
Method for using object-capturing device Download PDFInfo
- Publication number
- EP2685232B1 EP2685232B1 EP13004833.3A EP13004833A EP2685232B1 EP 2685232 B1 EP2685232 B1 EP 2685232B1 EP 13004833 A EP13004833 A EP 13004833A EP 2685232 B1 EP2685232 B1 EP 2685232B1
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- EP
- European Patent Office
- Prior art keywords
- capturing
- housing
- carrier
- capturing device
- microorganisms
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Not-in-force
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/02—Devices for withdrawing samples
- G01N1/22—Devices for withdrawing samples in the gaseous state
- G01N1/2273—Atmospheric sampling
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
- C12M41/36—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of biomass, e.g. colony counters or by turbidity measurements
Definitions
- the present invention relates to a method for using an object-capturing device, which captures objects, such as microorganisms and chemical substances, in the air.
- a well-known capturing device for capturing air-borne microorganisms has a carrier, which undergoes a phase transition from gel to sol at a temperature raised from room temperature, on a capturing dish, as disclosed in, for example, Japanese Patent Application Laid-Open No. 2009-131186 .
- Such capturing device is attached to an impactor-type air sampler.
- air sucked by the air sampler collides with the carrier, microorganisms carried by the air flow are captured by the carrier in a gel phase.
- the carrier solates by raising the temperature, and thereby the captured microorganisms with the carrier in a sol phase are obtained from the capturing dish.
- the obtained microorganisms are counted according to a predetermined counting method.
- a well-known method for counting microorganisms is the ATP method, which quantifies adenosine triphosphates (ATPs) extracted from microorganisms and thereby counts the microorganisms, as disclosed in, for example, Japanese Patent Application Laid-Open No. 11-137293 .
- the ATP method extracts ATPs contained in the microorganisms by contacting the captured microorganisms with an ATP extracting reagent, and counts the microorganisms based on the intensity of luminescence measured when the extracted ATPs reacts with a luminescence reagent.
- a method for counting captured microorganisms based on the number of microorganism colonies cultured in a plate medium requires several days.
- the ATP method requires about one to several hours from the capturing of microorganisms to the counting of the microorganisms.
- the ATP method dramatically reduces the required time.
- the ATP method counts microorganisms based on weak luminescence intensity. Substances that act as disturbance factors may be contained in a sample to be counted. Consequently, these substances need to be minimized.
- a conventional capturing device such as a device disclosed in Japanese Patent Application Laid-Open No. 2009-131186 , has a carrier exposed on a capturing dish.
- microorganisms other than those to be tested, or other substances that act as disturbance factors may attach onto the exposed carrier during the time after microorganisms are captured onto the carrier in an air sampler and before the microorganisms are counted.
- the carrier may be more heavily contaminated.
- the carrier is contaminated during the time after the carrier is removed from the air sampler and before the microorganisms are counted, and thereby the microorganisms captured at the test site may not be accurately counted.
- US 5 471 994 A discloses a cytology collection apparatus in which a body fluid to be analysed is captured by a porous arrangement which is encased between two detachable portions of a housing. Another such conventional apparatus is described in EP 0 636 873 A1 .
- An object of the present invention is to provide an object-capturing device using method which accurately detect objects such as microorganisms captured at a test site.
- the object-capturing device includes a capturing dish holding a carrier, which captures an object, on a first side of the capturing dish.
- the capturing dish has a through hole extending from the first side to a second side of the capturing dish.
- the present invention provides a method for using the object-capturing device as defined in present claim 1.
- the object-capturing device includes a carrier capturing an object, and a capturing dish holding the carrier on a first side of the capturing dish, and the capturing dish has a through hole extending from the first side to a second side of the capturing dish.
- the method includes the successive steps of: directing the carrier upward and capturing the object; and directing the carrier downward and contacting a reagent for detecting the object with the object captured on the carrier through the through hole.
- An object-capturing device to be used in an embodiment of the present invention will be described in detail with reference to the drawings as appropriate.
- the embodiment will be described using an object-capturing device for capturing air-borne microorganisms (for example, microbes and fungi) as an example.
- the objects captured by the object-capturing device using method of the present invention may be microscopic particles of metal or of chemical substances.
- the objects are not limited to solid objects, and may be mist.
- a microorganism counting apparatus 10 is an apparatus for counting microorganisms contained in a sample according to the ATP method.
- the microorganism counting apparatus 10 includes a mounting unit 102 mounting an object-capturing device 1 (refer to FIG. 2 ) having the sample therein, a functional liquid tank 105, a hot-water supplying unit 103, a suction unit 104, a reagent cartridge 2 having multiple reagents R, a dispensing unit 106, a luminescence-intensity measurement unit 107, and a control unit 108, which are housed in a cabinet 101.
- FIG. 1 shows the cabinet 101 and the reagent cartridge 2 in a dashed-two-dotted line, and omits the object-capturing device 1.
- the mounting unit 102 has a recessed portion 102a receiving the object-capturing device 1 (i.e., a housing 6).
- the mounting unit 102 also includes an engaging ring 102b.
- a heater 102c (refer to FIG. 3 ) is embedded in an aluminum member surrounding the recessed portion 102a, that is, forming the recessed portion 102a.
- other highly heat conductive members may be used instead of the aluminum member.
- the engaging ring 102b is attached on the periphery of the opening of the recessed portion 102a.
- the housing 6 is mounted on the mounting unit 102 by engaging the engaging ring 102b with first engaging claws 62a provided on the housing 6 of the object-capturing device 1.
- the engaging ring 102b has cutout portions 102d in such a planar shape as to receive respective first engaging claws 62a of the housing 6.
- a gap G is formed to have a height large enough for receiving the first engaging claws 62a.
- the housing 6 when the housing 6 is fitted into the recessed portion 102a, the first engaging claws 62a are inserted into the recessed portion 102a through respective cutout portions 102d, and the housing 6 is rotated to slide the first engaging claws 62a into the gap G. Thereby, the housing 6 is engaged with the engaging ring 102b.
- FIG. 3 a through hole 41 for injecting the reagents R (refer to FIG. 1 ) and hot water into an internal space 66 of the housing 6, a carrier 5 disposed on the back side of the capturing dish 4, a discharge opening 64a of the housing 6, a filter 7 provided on the outside of the discharge opening 64a, and a suction head 104a of the suction unit 104 (refer to FIG. 1 ) connected with the housing 6 are shown.
- the internal space 66 corresponds to "a space" in the claims.
- the object-capturing device 1 is mounted on the mounting unit 102 through the engaging ring 102b.
- the capturing dish 4 having the through hole 41 is disposed above the housing 6, and the carrier 5 as a sample is disposed on the back side of the capturing dish 4 and in the internal space 66 of the housing 6.
- the housing 6 has a protruding portion which protrudes upward from the recessed portion 102a, and the capturing dish 4 is disposed in the protruding portion.
- the heater 102c may be any means that is capable of heating the recessed portion 102a (the aluminum member), which surrounds the housing 6 of the object-capturing device 1 mounted on the mounting unit 102, to a predetermined temperature.
- the heater 102c is preferable to be, for example, a cartridge heater.
- the functional liquid tank 105 shown in FIG. 1 is adapted to store liquid such as sterile distilled water. This liquid is poured into the housing 6 (refer to FIG. 3 ) to, for example, improve a filtration rate of the carrier 5 (refer to FIG. 3 ) of the object-capturing device 1, or to wash the housing, as described below. The liquid is also poured into a piping system connected to a syringe pump 106c of the dispensing unit 106 as described below.
- the functional liquid tank 105 may store buffer solution.
- the hot-water supplying unit 103 heats and supplies, for example, the sterile distilled water supplied from the functional liquid tank 105. Specifically, the hot-water supplying unit 103 injects hot water into the internal space 66 (refer to FIG. 3 ) in the housing 6 through the through hole 41 (refer to FIG. 3 ) of the capturing dish 4.
- the hot-water supplying unit 103 is a unit (not shown) for discharging hot water heated by a cartridge heater with a peristaltic pump through a nozzle formed with, for example, a flexible tube. This nozzle having a means for moving in horizontal and vertical directions is inserted into the internal space 66 (refer to FIG. 3 ) of the housing 6 through the through hole 41 (refer to FIG. 3 ) of the capturing dish 4 as necessary.
- the suction unit 104 shown in FIG. 1 sucks, for example, the hot water and the reagents R described below, which are injected into the internal space 66 (refer to FIG. 3 ) in the housing 6, to discharge them through the filter 7 (refer to FIG. 3 ).
- this suction unit 104 includes the suction head 104a (refer to FIG. 3 ) described above, a suction pump (not shown) connected with the suction head 104a through predetermined piping, and a waste tank.
- the suction unit 104 to be used according to the embodiment further includes a lifting apparatus (not shown) lifting and lowering the suction head 104a to enable the suction head 104a (refer to FIG. 3 ) to be engaged with and disengaged from the housing 6 (refer to FIG. 3 ) mounted on the mounting unit 102.
- a lifting apparatus (not shown) lifting and lowering the suction head 104a to enable the suction head 104a (refer to FIG. 3 ) to be engaged with and disengaged from the housing 6 (refer to FIG. 3 ) mounted on the mounting unit 102.
- each reagent R is disposed at a predetermined position in the vicinity of the mounting unit 102.
- each reagent R is disposed at a predetermined position, and a dispensing nozzle 106a of the dispensing unit 106 described below dispenses the reagents R in a predetermined order into the housing 6 of the object-capturing device 1 and into a luminescence-test tube 107a of the luminescence-intensity measurement unit 107.
- the location (the coordinates) of each reagent R is stored in the control unit 108 controlling the dispensing unit 106, as described below.
- Examples of the reagents R necessary to the ATP method includes an ATP eliminating reagent for eliminating ATPs that exist out of cells of captured microorganisms, an ATP extracting reagent for extracting ATPs contained in the microorganisms, and an ATP luminescence reagent for producing luminescence of the ATPs extracted from the microorganisms.
- Examples of the ATP eliminating reagent include an ATP-degrading enzyme.
- ATP extracting reagent examples include a benzalkonium chloride, a trichloroacetic acid, and a Tris buffer solution.
- ATP luminescence reagent examples include a luciferase/luciferin reagent.
- the examples of the reagents R may include a correction reagent for the luminescence-intensity measurement unit 107, and sterile pure water.
- the dispensing unit 106 shown in FIG. 1 dispenses the reagents R described above into the housing 6 of the object-capturing device 1.
- the dispensing unit 106 also dispenses the reagents R and ATP extracted solution in the housing 6 (refer to FIG. 3 ) as described below into the luminescence-test tube 107a of the luminescence-intensity measurement unit 107.
- the dispensing unit 106 may include the dispensing nozzle 106a formed with a thin tube, an actuator 106b moving the dispensing nozzle 106a in the xyz axis directions, the syringe pump 106c connected with the dispensing nozzle 106a through predetermined flexible piping, the piping (not shown) supplying, for example, sterile distilled water from the functional liquid tank 105 through the syringe pump 106c to the dispensing nozzle 106a.
- the luminescence-intensity measurement unit 107 shown in FIG. 1 may be a unit which includes the luminescence-test tube 107a for receiving the ATP extracted solution dispensed from the housing 6 (refer to FIG. 3 ) as described below to produce luminescence of ATPs, and a luminescence detecting unit body 107b having, for example, a photomultiplier which detects luminescence intensity of the ATPs.
- the control unit 108 shown in FIG. 1 has overall control over the microorganism counting apparatus 10.
- the control unit 108 also controls the hot-water supplying unit 103, the suction unit 104, the dispensing unit 106, and the luminescence-intensity measurement unit 107 according to a procedure to be described below, after the object-capturing device 1 (refer to FIG. 3 ) is mounted on the mounting unit 102.
- This control unit 108 includes a CPU, a ROM, a RAM, various interfaces, and circuitry.
- FIG. 4 which is referred to in the following description, is a flowchart showing the procedure in which the microorganism counting apparatus operates based on instructions of the control unit.
- control unit 108 starts execution of the following procedure when a start switch (not shown) is turned on after the object-capturing device 1 (refer to FIG. 3 ) is mounted on the mounting unit 102.
- the control unit 108 sends an instruction to, for example, a predetermined inverter to apply power to the heater 102c (refer to FIG. 3 ) to generate heat.
- a predetermined inverter to apply power to the heater 102c (refer to FIG. 3 ) to generate heat.
- the temperature of the carrier 5 (refer to FIG. 3 ) of the object-capturing device 1 is raised with the heater 102c (Step S201).
- the carrier 5 solates and falls down off the capturing dish 4 (refer to FIG. 3 ) onto the inside bottom of the housing 6 (refer to FIG. 3 ).
- the control unit 108 sends an instruction to the hot-water supplying unit 103 (refer to FIG. 1 ) to inject hot water into the housing 6 (refer to FIG. 3 ) (Step S202). Thereby, the carrier 5 (refer to FIG. 3 ) further solates, and is diluted with the hot water.
- the control unit 108 sends an instruction to the suction unit 104 (refer to FIG. 1 ) to engage the suction head 104a (refer to FIG. 3 ) with the housing 6 (refer to FIG. 3 ), and then suck and filter the contents in the housing 6 (refer to FIG. 3 ) (Step S203). Thereby, the microorganisms captured in the carrier 5 are separated and held by the filter 7 (refer to FIG. 3 ), and the diluted carrier 5 is filtered and discharged out of the housing 6.
- the control unit 108 again sends an instruction to the hot-water supplying unit 103 to dispense hot water into the housing 6 (refer to FIG. 3 ) (Step S204). Then, the hot water in the housing 6 is filtered again (Step S205). Thereby, the filtered hot water removes remaining diluted carrier 5 from the inside of the housing 6, and accordingly the recovery rate of microorganisms at the filter 7 is improved.
- the control unit 108 sends to an instruction to the dispensing unit 106 (refer to FIG. 1 ) to dispense the ATP eliminating reagent in the reagent cartridge 2 into the housing 6 (refer to FIG. 3 ) (Step S206).
- the ATPs that exist out of the cells of the microorganisms on the filter 7 are eliminated.
- the control unit 108 sends an instruction to the suction unit 104 (refer to FIG. 1 ) to suck the contents of the housing 6 (refer to FIG. 3 ) and filter the sucked contents (Step S207). Thereby, the microorganisms are separated and held by the filter 7 (refer to FIG. 3 ), and the ATP eliminating reagent and the hot water are filtered and discharged out of the housing 6.
- the control unit 108 sends an instruction to the dispensing unit 106 (refer to FIG. 1 ) to dispense the ATP luminescence reagent in the reagent cartridge 2 into the luminescence-test tube 107a (refer to FIG. 1 ) (Step S208).
- the control unit 108 sends an instruction to the luminescence-intensity measurement unit 107 (refer to FIG. 1 ) to turn on the luminescence detecting unit body 107b (refer to FIG. 1 ) (Step S209).
- the control unit 108 sends an instruction to the dispensing unit 106 (refer to FIG. 1 ) to dispense the ATP extracting reagent in the reagent cartridge 2 into the housing 6 (refer to FIG. 3 ) (Step S210). Thereby, ATPs are extracted from the microorganisms held by the filter 7, and sample solution is prepared on the filter 7.
- the luminescence detecting unit of the luminescence-intensity measurement unit 107 performs a background measurement of the luminescence-test tube 107a with the ATP luminescence reagent.
- the control unit 108 sends an instruction to the dispensing unit 106 (refer to FIG. 1 ) to dispense an adequate amount of the sample solution (i.e., the ATP extracted solution) in the housing 6 into the luminescence-test tube 107a where the background measurement has been performed (Step S211).
- the ATP extracted solution reacts with the ATP luminescence reagent which has been dispensed in Step S208, and produces luminescence in the luminescence-test tube 107a.
- the luminescence detecting unit body 107b (refer to FIG. 1 ) of the luminescence-intensity measurement unit 107 (refer to FIG. 1 ) detects the ATP luminescence and outputs signals.
- the control unit 108 digitizes the outputted signals, and measures luminescence intensity based on the single-photon counting method (Step S212).
- control unit 108 calculates the ATP amount (amol) in the ATP extracted solution dispensed into the luminescence-test tube 107a based on a prestored calibration curve indicating the relation between the ATP amount (amol) and the luminescence intensity (CPS), and the control unit 108 counts the microorganisms using an ATP value, which may be converted into the equivalent number of the microorganisms in the carrier 5.
- This ATP value is calculated based on the ATP amount (amol) and the amount of the ATP extracted solution of the sample solution prepared in Step S210 (Step S213).
- FIG. 5 is a perspective view showing the object-capturing device to be used according to the embodiment.
- FIG. 6A is an exploded perspective view showing the object-capturing device of FIG. 5 viewed from obliquely above.
- FIG. 6B is an exploded perspective view showing the object-capturing device of FIG. 5 viewed from obliquely below.
- FIG. 7 is a cross-sectional view along a line VII-VII in FIG. 5 .
- the object-capturing device 1 to be used according to the embodiment is placed in an impactor-type air sampler 50 (refer to FIG. 8 ) described below to capture microorganisms, which are air-borne objects.
- the object-capturing device 1 is mounted in the microorganism counting apparatus 10 described above to count the captured microorganism.
- the object-capturing device 1 is turned upside down and used when microorganisms are captured, as described in detail below.
- an upper portion of the object-capturing device 1 is in a substantially cylindrical shape, and a lower portion of the object-capturing device 1 is in a substantially conical shape so that the diameter of the horizontal cross section of the object-capturing device 1 becomes small downward as the horizontal cross section lowers.
- the object-capturing device 1 is engaged with the air sampler 50 (refer to FIG. 8 ) in the upper portion of the object-capturing device 1 when microorganisms are captured.
- the object-capturing device 1 is engaged with the microorganism counting apparatus 10 in the lower portion of the object-capturing device 1 when the captured microorganisms are counted.
- FIG. 5 the cover 3, the housing 6, second engaging claws 31 engaging with the air sampler 50 (refer to FIG. 8 ) described below, and the first engaging claws 62a engaging with the microorganism counting apparatus 10 are shown.
- the air sampler 50 corresponds to "a capturing apparatus which captures the object" in the claims.
- Each second engaging claw 31 corresponds to "an engaging portion" in the claims.
- the object-capturing device 1 includes the cover 3, the capturing dish 4, the carrier 5, the housing 6, the filter 7, and a filter-securing ring 8, which are disposed in this order from upward to downward and fitted with each other.
- the cover 3 is attached to close an upper opening of the housing 6 described below, and has a cylindrical shape with a bottom and an opening facing upward.
- the second engaging claws 31 described above are formed to protrude radially outward, and to be disposed in a constant spacing with each other on the circumferential surface of the cover 3.
- the number of the second engaging claws 31 is three, which is same as the number of engaging cutout portions 53 (refer to FIG. 8 ) described below of the air sampler 50.
- three third engaging claws 32 are formed to protrude radially outward, and to be disposed in a constant spacing with each other on the circumferential surface of the cover 3.
- the third engaging claws 32 are fitted in respective first L-shaped grooves 61a described below of the housing 6 to detachably engage the cover 3 with the housing 6.
- an outer bottom surface of the cover 3 forms an uneven surface constituted by multiple straight ridges and straight grooves disposed alternately and in parallel with each other, the ridges of which protrude downward.
- this uneven outer bottom surface reduces the area of contact with the capturing dish 4.
- the object-capturing device 1 when the object-capturing device 1 is carried to a microorganism counting facility (for example, a facility having the microorganism counting apparatus 10 (refer to FIG. 1 )) at low temperature as necessary, condensation may rarely occur between the cover 3 and the capturing dish 4. Even in this case, the uneven surface facilitates easy detachment of the cover 3 from the capturing dish 4.
- This uneven surface is not limited to the above straight ridges and straight grooves, and may be formed with multiple protrusions, or with grains such as a matte finish pattern or a texture pattern.
- a protrusion 33 in a cylindrical shape is formed to protrude downward.
- the protrusion 33 has an outer diameter rather smaller than the inner diameter of the through hole 41 of the capturing dish 4 to be described below.
- the height of the protrusion 33 equals to that of the through hole 41.
- the capturing dish 4 has a disk shape.
- the through hole 41 which is bored through the capturing dish 4 in the vertical direction, is formed in a center portion of the capturing dish 4.
- the upper surface of the capturing dish 4 forms an even surface to be made in contact with the outer bottom surface of the cover 3 described above.
- double ring-shaped ribs 42a and 42b extend vertically on the periphery of the through hole 41 in an inner portion and an outer portion, to receive the ring-shaped carrier 5 as described below.
- the outer diameter of the capturing dish 4 ranges between the inner diameter of a lower cylinder portion 62 and the inner diameter of an upper cylinder portion 61 of the housing 6 (including both end values).
- the outer diameter of the capturing dish 4 is substantially equal to the inner diameter of the upper cylinder portion 61.
- the outer diameter of the ring-shaped rib 42b formed in the outer portion of the capturing dish 4 shown in FIG. 6B is equal to the inner diameter of the lower cylinder portion 62 described below, or less.
- the outer diameter of the ring-shaped rib 42b is substantially equal to the inner diameter of the lower cylinder portion 62.
- the carrier 5 is placed in the air sampler 50 (refer to FIG. 8 ), as described below, to receive air flow when the air sampler 50 sucks the air, and to capture microorganisms carried in the air flow.
- the carrier 5 is made of a material that undergoes a phase transition from gel to sol when the temperature rises from the room temperature.
- the material of the carrier 5 is preferably such a material as to undergoes a phase transition from gel to sol at 30°C or higher. More preferably, the material liquefies at a temperature between 37°C and 40°C.
- the material is a gelatin, a mixture of gelatin and glycerol, or a copolymer having a ratio of N-acryloylglycinamide to N-methacryloyl-N'-biotinyl propylene diamine of 10:1.
- the carrier 5 has a ring shape, as described above. As shown in FIG. 6B , the carrier 5 preferably has the same shape as that of the space formed between the ring-shaped ribs 42a and 42b of the capturing dish 4.
- the carrier 5 may be formed by applying the material described above to the space formed between the ring-shaped ribs 42a and 42b, or by filling the space with the material. Instead, a separated ring-shaped carrier may be fitted into the space.
- the housing 6 has the upper cylinder portion 61 having the inner diameter substantially as large as the outer diameter of the cover 3 as described above, the lower cylinder portion 62 having the inner diameter smaller than the inner diameter of the upper cylinder portion 61, a conical portion 64 being in an inverted conical shape with an inner diameter which becomes smaller from the inner diameter of the lower cylinder portion 62, and a filter fitting portion 65 provided on the periphery of an outlet of the discharge opening 64a formed in the lowest portion of the conical portion 64, which are disposed in this order from upward to downward to form an integral unit.
- the lower cylinder portion 62 is connected with the upper cylinder portion 61 through a shelf portion 63.
- the first engaging claws 62a are formed to be engaged with engaging ring 102b (refer to FIG. 2 ) of the microorganism counting apparatus 10 described above.
- the first engaging claws 62a protrude outward in the radial direction of the lower cylinder portion 62, and are disposed in a constant spacing with each other on the circumferential surface of the lower cylinder portion 62.
- the number of the first engaging claws 62a is four.
- the conical portion 64 having the inner diameter becoming smaller downward enables the contents to easily flow down toward the lowest portion, that is, the discharge opening 64a (refer to FIG. 6B ).
- the filter fitting portion 65 forms an integral unit with a filter housing portion 65a (refer to FIG. 6B ) forming a thin disk-shaped space, the shape of which matches that of the filter 7 which is disposed to close the outlet of the discharge opening 64a (refer to FIG. 6B ), and a ring supporting portion 65b having a cylindrical shape and supporting the filter-securing ring 8.
- Second L-shaped grooves 65c are formed on the inner circumferential surface of the ring supporting portion 65b, and fourth engaging claws 82a formed on the filter-securing ring 8 described below are fitted into respective second L-shaped grooves 65c.
- the number of the second L-shaped grooves 65c is four, and the second L-shaped grooves 65c are formed to be disposed in a constant spacing with each other on the circumferential surface of the ring supporting portion 65b.
- the filter 7 to be used according to the embodiment is a membrane filter. As described above, the filter 7 closes the outlet of the discharge opening 64a. In other words, the filter 7 is disposed on the outside of the discharge opening 64a.
- the filter 7 includes a hydrophilic filter 7a and a hydrophobic filter 7b, which are arranged in this order from the discharge opening 64a.
- the hydrophilic filter 7a and the hydrophobic filter 7b may be selected from products launched in the market.
- Examples of the hydrophilic filter 7a include MF-Millipore (Nihon Millipore K.K.), Durapore (Nihon Millipore K.K.), and Isopore (Nihon Millipore K.K.).
- hydrophobic filter 7b examples include Mitex (Nihon Millipore K.K.) and Polypropylene Prefilter (Nihon Millipore K.K.).
- the filter 7 used in the embodiment should have an outer diameter larger than the inner diameter of the discharge opening 64a.
- the filter-securing ring 8 fixes the filter 7 to the housing 6 (i.e., the conical portion 64).
- the filter-securing ring 8 has a through hole 81 at a position where the through hole 81 communicates with the discharge opening 64a of the conical portion 64 through the filter 7.
- the filter-securing ring 8 includes a ring body 82 having a shape substantially same as the inner diameter of the ring supporting portion 65b of the filter fitting portion 65 described above, and a flange portion 83 formed on the lower side of the ring body 82 and having a diameter larger than the outer diameter of the ring body 82.
- the filter-securing ring 8 further includes: a fitting portion 84 which is deposited on the ring body 82 so that the fitting portion 84 and the ring body 82 form an integral unit, and is fitted into the filter housing portion 65a of the housing 6; and a ring-shaped rib 85 vertically disposed on the periphery of an opening of the through hole 81 of the fitting portion 84.
- the ring-shaped rib 85 presses the filter 7 on the periphery of the outlet of the discharge opening 64a (refer to FIG. 6B ).
- the fourth engaging claws 82a are formed to protrude radially outward, and to be disposed in a constant spacing with each other on the circumferential surface of the ring body 82.
- the fourth engaging claws 82a are formed at positions corresponding to the respective second L-shaped grooves 65c of the ring supporting portion 65b described above, and are fitted into the respective second L-shaped grooves 65c to detachably engage the filter-securing ring 8 with the housing 6.
- the object-capturing device 1 as described above is formed as follows.
- the capturing dish 4 is mounted on the shelf portion 63 of the housing 6; the housing 6 is coupled to the cover 3 through the capturing dish 4 using the first L-shaped grooves 61a and the third engaging claws 32; and the through hole 41 of the capturing dish 4 is sealed by the protrusion 33 of the cover 3.
- the housing 6 is decoupled from the cover 3 by rotating the housing 6 relative to the cover 3 to disengage the third engaging claws 32 from the first L-shaped grooves 61a.
- the filter 7 is disposed in the filter housing portion 65a to close the outlet of the discharge opening 64a of the conical portion 64, and the filter fitting portion 65 is engaged with the filter-securing ring 8 using the second L-shaped grooves 65c and the fourth engaging claws 82a described above. Thereby, the discharge opening 64a of the conical portion 64 communicates with the through hole 81 of the filter-securing ring 8 through the filter 7.
- the filter fitting portion 65 is engaged with the filter-securing ring 8
- the filter 7 is pressed by the ring-shaped rib 85 of the filter-securing ring 8, and thereby the filter 7 is disposed on the periphery of the outlet of the discharge opening 64a.
- the filter 7 is fixed firmly.
- the object-capturing device 1 as described above is used as follows.
- the through hole 41 opens to communicate with the internal space 66 which receives the reagents, as shown in FIG. 3 .
- the protrusion 33 of the cover 3 seals the through hole 41.
- the outlet of the discharge opening 64a of the conical portion 64 is closed by the filter 7 which separates microorganisms.
- the internal space 66 is a space isolated from the external environment (i.e., a closed space) at least for microorganisms. Consequently, the carrier 5 held on the capturing dish 4 is placed in this closed space.
- the object-capturing device 1 as described above other than the filter 7 may be molded with resin, preferably polypropylene.
- FIG. 8 referred to below is a perspective view showing the method for capturing microorganisms using the object-capturing device of the present invention.
- the object-capturing device 1 when microorganisms are captured, the object-capturing device 1 is used in such a way that the capturing dish 4 holding the carrier 5 is mounted on the cover 3.
- the object-capturing device 1 shown in FIG. 7 is turned upside down and used with the capturing dish 4 left on the cover 3 and with the housing 6 and the filter-securing ring 8 removed.
- the housing 6 is removed from the cover 3 by rotating the housing 6 relative to the cover 3 to disengage the third engaging claws 32 (refer to FIG. 6A ) from the first L-shaped grooves 61a (refer to FIG. 6A ) as described above after the cover 3 is located in the pedestal 52 of the air sampler 50 as described below.
- the object-capturing device 1 is mounted on the pedestal 52 in a round shape in plan view, which is formed on the upper side of an air sampler body 51 of the air sampler 50.
- the pedestal 52 has the engaging cutout portions 53 formed to receive the second engaging claws 31 of the cover 3, and thereby the object-capturing device 1 is located in a center portion of the pedestal 52.
- FIG. 8 shows suction openings 54 of the air sampler body 51, and a nozzle head 55 of the air sampler 50.
- the housing 6 and the filter-securing ring 8 which form an integral unit are removed to expose the carrier 5 of the object-capturing device 1 mounted in the pedestal 52, and the nozzle head 55 is placed over the pedestal 52, as shown in FIG. 8 .
- the process of exposing the carrier 5 corresponds to "the step of exposing the carrier" in the claims.
- a fan (not shown) disposed in the air sampler body 51 is activated, and the air is sucked through the suction openings 54. Then, air flow is injected to the carrier 5 from multiple micro nozzles (not shown) provided in the nozzle head 55. As a result, microorganisms carried in the air injected to the carrier 5 are captured by the carrier 5. In other words, microorganisms are captured with the carrier 5 directed upward.
- the protrusion 33 of the cover 3 seals the through hole 41 (refer to FIG. 7 ) of the capturing dish 4.
- the surface of the capturing dish 4 on the side of the carrier 5 is flush with the bottom of the protrusion 33. This reduces disturbance of the received air flow. Consequently, the carrier 5 is capable of capturing microorganisms efficiently.
- This capturing process corresponds to "the step of capturing the object" in the claims.
- the object-capturing device 1 as shown in FIG. 7 is carried by a user into the microorganism counting apparatus 10, as described above (refer to FIG. 2 ).
- the object-capturing device 1 is mounted on the mounting unit 102, and the cover 3 is removed from the housing 6 by the user (refer to FIG. 3 ).
- FIGs 9A1 to 9A4 are cross-sectional views of the object-capturing device, showing the method for using the object-capturing device in the microorganism counting apparatus.
- FIGs 9B1 to 9B4 are enlarged schematic diagrams showing the vicinity of the filter in the case of FIGs 9A1 to 9A4 .
- FIGs 9B1 to 9B4 show microorganisms B and ATPs. However, the sizes of actual microorganisms are as small as in the order of micrometer, and the sizes of actual ATPs are as small as that of a molecule. Accordingly, FIGs 9B1 to 9B4 shows no relative sizes of a microorganism and an ATP.
- Step S201 When the temperature of the carrier 5 is raised in Step S201 (refer to FIG. 4 ) as described above, the carrier 5 held on the capturing dish 4 solates and falls down onto the conical portion 64 of the housing 6 as shown in FIG. 9A1 .
- the microorganisms B captured with the air sampler 50 (refer to FIG. 8 ) are retained with the carrier 5 on the filter 7 as shown in FIG. 9B1 .
- the carrier 5 When the hot water HW is injected into the housing 6 in Step S202 (refer to FIG. 4 ) as described above, the carrier 5 further solates and is diluted by the hot water.
- the filter 7 includes the hydrophobic filter 7b on the lower side thereof as shown in FIG. 9A2 .
- the hot water HW containing the diluted carrier 5 is retained in the housing 6.
- the microorganisms B are retained in the hot water HW containing the diluted carrier 5 on the filter 7.
- FIG. 9A2 also shows the capturing dish 4, and the conical portion 64 (The same reference number denotes the same element throughout the following views.)
- Step S203 When the contents in the housing 6 are filtered in Step S203 (refer to FIG. 4 ) as described above, the hot water HW containing the diluted carrier 5 in the housing 6 (refer to FIG. 9A2 ) is discharged as shown in FIG. 9A3 .
- the microorganisms B in the hot water HW containing the diluted carrier 5 are separated and held by the filter 7 as shown in FIG. 9B3 .
- the filter 7 to be used according to the embodiment has a double layer structure of the hydrophilic filter 7a and the hydrophobic filter 7b.
- the hydrophobic filter 7b enables liquid to be retained on the filter unless the liquid is sucked or pressure-filtered. This enables reaction with reagent, such as ATP extraction, to be performed on the filter 7.
- the ATP eliminating reagent is dispensed into the housing 6 in Step S206 (refer to FIG. 4 ) as described above, and then the ATP extracting reagent is dispensed into the housing 6 in Step S210 (refer to FIG. 4 ) as described above.
- ATP extracted solution EX is retained as shown in FIG. 9A4 .
- the ATP extracted solution EX contains ATPs, the amount of which corresponds to the number of the microorganisms B.
- the ATP extracted solution EX shown in FIG. 9B4 is dispensed into the luminescence-test tube 107a (refer to FIG. 1 ) in Step S211 (refer to FIG. 4 ) as described above, and then the process sequence of the method for using the object-capturing device 1 ends.
- microorganisms are captured with the carrier 5 directed upward, and then the carrier 5 is directed downward to contact the microorganisms with the reagents through the through hole 41.
- the carrier 5 directed upward facilitates the capturing of the microorganisms.
- the carrier 5 is directed downward, and thereby the capturing dish 4 serves as a cover of the carrier 5. For example, this prevents the carrier 5 from being contaminated with, for example, dust and microbes falling from above.
- the carrier 5 is placed in the closed space in the housing 6.
- the carrier 5 is prevented from being contaminated with substances which are disturbance factors for the counting of the microorganisms, unlike a conventional capturing device with an exposed carrier, such as the device disclosed in Japanese Patent Application Laid-Open No. 2009-131186 .
- the object-capturing device 1 and the using method thereof enable more accurate counting of the microorganisms captured at a test site.
- the object-capturing device 1 and the using method thereof when the microorganisms are contacted with the reagents R using the microorganism counting apparatus 10, the reagents R are dispensed into the housing 6 through the through hole 41 of the capturing dish 4. This minimizes the communication between the inside and the outside of the housing 6. As a result, the inside of the housing 6 is prevented from being contaminated with substances which are disturbance factors for the counting of the microorganisms.
- the housing 6 has the discharge opening 64a through which the contents are discharged, and the discharge opening 64a has the filter disposed thereon to separate and hold microorganisms.
- the microorganisms may be contacted with the reagents R in the housing 6. Consequently, the object-capturing device 1 and the using method thereof dramatically reduce the disturbance factors for the counting of the microorganisms, unlike a conventional capturing device, such as the device disclosed in Japanese Patent Application Laid-Open No. 2009-131186 , which is used in a way such that microorganisms are extracted from the capturing device and the extracted microorganisms are contacted with reagents for counting.
- the filter 7 has a double layer structure of the hydrophilic filter 7a and the hydrophobic filter 7b. Thereby, reaction of reagents with recovered microorganisms may be performed on the filter 7, unlike a filter used in the conventional ATP methods, which includes only a hydrophilic filter.
- the cover 3 has the second engaging claws 31 formed on the opposite side of the housing 6 to engage with the air sampler.
- the housing 6, which is fixed with the cover 3 to form an integral unit as shown in FIG. 5 is grasped by hands, the cover 3 is placed into the air sampler 50 as shown in FIG. 8 , and then the housing 6 is removed from the cover 3.
- the capturing dish 4 holding the carrier 5 is not touched by hands. Consequently, the object-capturing device 1 and the using method thereof surely prevent the carrier 5 from being contaminated with substances which are disturbance factors for the counting of the microorganism.
- the object-capturing device 1 According to the object-capturing device 1 and the using method thereof, after the object-capturing device 1 is mounted on the mounting unit 102, when the cover 3 is removed from the housing 6 by a user, the capturing dish 4 is turned upside down relative to the state at the time when placed in the air sampler, and thereby the carrier 5 faces toward the internal space 66. This surely prevents the contamination of the carrier 5.
- This advantage may be achieved regardless of the type or the number of the filter 7, such as the hydrophilic filter 7a or the hydrophobic filter 7b described above.
- the microorganisms captured with the object-capturing device 1 are counted in the microorganism counting apparatus 10.
- the reagents R may be manually dispensed into the housing to count the microorganisms by the ATP method.
- the present invention is applicable to spore-forming bacteria such as Bacillus subtilis.
- examples of the reagents described above may include a vegetative cell conversion reagent, such as amino acid and sugar.
- the ATP method is used to count microorganisms.
- the microorganisms may be counted based on the fluorescence produced when substances in a living body, such as DNA, RNA, and NAD, which are extracted from the microorganisms, are irradiated with excitation light.
- the counting may be made based on endotoxin contained in the cell membrane of gram negative bacilli.
- microbes may be counted based on luminescence intensity resulting from the limulus test on the endotoxin.
- the microorganisms may be counted by recovering the microorganisms from the filter 7 and culturing the recovered microorganisms.
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Description
- The present invention relates to a method for using an object-capturing device, which captures objects, such as microorganisms and chemical substances, in the air.
- Conventionally, a technique of capturing objects, such as air-borne microorganisms and chemical substances, by sucking air through a filter and separating the objects by the filter has been well-known. A well-known capturing device for capturing air-borne microorganisms has a carrier, which undergoes a phase transition from gel to sol at a temperature raised from room temperature, on a capturing dish, as disclosed in, for example, Japanese Patent Application Laid-Open No.
2009-131186 - A well-known method for counting microorganisms is the ATP method, which quantifies adenosine triphosphates (ATPs) extracted from microorganisms and thereby counts the microorganisms, as disclosed in, for example, Japanese Patent Application Laid-Open No.
11-137293 - For example, a method for counting captured microorganisms based on the number of microorganism colonies cultured in a plate medium requires several days. On the other hand, the ATP method requires about one to several hours from the capturing of microorganisms to the counting of the microorganisms. Thus, the ATP method dramatically reduces the required time.
- However, the ATP method counts microorganisms based on weak luminescence intensity. Substances that act as disturbance factors may be contained in a sample to be counted. Consequently, these substances need to be minimized.
- A conventional capturing device , such as a device disclosed in Japanese Patent Application Laid-Open No.
2009-131186 - In other words, when the capturing device, such as the device disclosed in Japanese Patent Application Laid-Open No.
2009-131186 -
US 5 471 994 A discloses a cytology collection apparatus in which a body fluid to be analysed is captured by a porous arrangement which is encased between two detachable portions of a housing. Another such conventional apparatus is described inEP 0 636 873 A1 . - An object of the present invention is to provide an object-capturing device using method which accurately detect objects such as microorganisms captured at a test site.
- To solve the above problems, the object-capturing device includes a capturing dish holding a carrier, which captures an object, on a first side of the capturing dish. The capturing dish has a through hole extending from the first side to a second side of the capturing dish.
- To solve the above problems, the present invention provides a method for using the object-capturing device as defined in
present claim 1. The object-capturing device includes a carrier capturing an object, and a capturing dish holding the carrier on a first side of the capturing dish, and the capturing dish has a through hole extending from the first side to a second side of the capturing dish. In an example useful to understand the invention, the method includes the successive steps of: directing the carrier upward and capturing the object; and directing the carrier downward and contacting a reagent for detecting the object with the object captured on the carrier through the through hole. -
-
FIG. 1 is a diagram showing a structure of a microorganism counting apparatus having an object-capturing device mounted therein to be used in an embodiment of the present invention; -
FIG. 2 is a perspective view showing the vicinity of a mounting unit for the object-capturing device in the microorganism counting apparatus ofFIG. 1 ; -
FIG. 3 is a cross-sectional view showing the object-capturing device mounted on the mounting unit in the microorganism counting apparatus ofFIG. 1 ; -
FIG. 4 is a flowchart of operations of the microorganism counting apparatus based on instructions of a control unit; -
FIG. 5 is a perspective view showing the object-capturing device to be used in the embodiment; -
FIG. 6A is an exploded perspective view showing the object-capturing device ofFIG. 5 viewed from obliquely above; -
FIG. 6B is an exploded perspective view showing the object-capturing device ofFIG. 5 viewed from obliquely below; -
FIG. 7 is a cross-sectional view along a line VII-VII inFIG. 5 ; -
FIG. 8 is a perspective view showing a method for capturing microorganisms using the object-capturing device according to the embodiment of the present invention; -
FIG. 9A1 is a cross-sectional view of the object-capturing device, showing a method for using the object-capturing device in the microorganism counting apparatus; -
FIG. 9A2 is a cross-sectional view of the object-capturing device, showing the method for using the object-capturing device in the microorganism counting apparatus; -
FIG. 9A3 is a cross-sectional view of the object-capturing device, showing the method for using the object-capturing device in the microorganism counting apparatus; -
FIG. 9A4 is a cross-sectional view of the object-capturing device, showing the method for using the object-capturing device in the microorganism counting apparatus; -
FIG. 9B1 is an enlarged schematic diagram showing the vicinity of a filter in the case ofFIG. 9A1 ; -
FIG. 9B2 is an enlarged schematic diagram showing the vicinity of the filter in the case ofFIG. 9A2 ; -
FIG. 9B3 is an enlarged schematic diagram showing the vicinity of the filter in the case ofFIG. 9A3 ; and -
FIG. 9B4 is an enlarged schematic diagram showing the vicinity of the filter in the case ofFIG. 9A4 . - An object-capturing device to be used in an embodiment of the present invention will be described in detail with reference to the drawings as appropriate. The embodiment will be described using an object-capturing device for capturing air-borne microorganisms (for example, microbes and fungi) as an example. However, the objects captured by the object-capturing device using method of the present invention may be microscopic particles of metal or of chemical substances. The objects are not limited to solid objects, and may be mist.
- First, an overall structure of a microorganism counting apparatus having the object-capturing device mounted therein, and a method for counting microorganisms with the microorganism counting apparatus will be described according to the embodiment. Second, the object-capturing device and a method for using the object-capturing device will be described according to the embodiment.
- As shown in
FIG. 1 , amicroorganism counting apparatus 10 is an apparatus for counting microorganisms contained in a sample according to the ATP method. Themicroorganism counting apparatus 10 includes a mountingunit 102 mounting an object-capturing device 1 (refer toFIG. 2 ) having the sample therein, afunctional liquid tank 105, a hot-water supplying unit 103, asuction unit 104, a reagent cartridge 2 having multiple reagents R, adispensing unit 106, a luminescence-intensity measurement unit 107, and acontrol unit 108, which are housed in acabinet 101. - For simplicity,
FIG. 1 shows thecabinet 101 and the reagent cartridge 2 in a dashed-two-dotted line, and omits the object-capturingdevice 1. - As shown in
FIG. 2 , the mountingunit 102 has a recessedportion 102a receiving the object-capturing device 1 (i.e., a housing 6). The mountingunit 102 also includes anengaging ring 102b. As described below, aheater 102c (refer toFIG. 3 ) is embedded in an aluminum member surrounding the recessedportion 102a, that is, forming the recessedportion 102a. To form the recessedportion 102a, other highly heat conductive members may be used instead of the aluminum member. - The engaging
ring 102b is attached on the periphery of the opening of the recessedportion 102a. As described in detail below, thehousing 6 is mounted on the mountingunit 102 by engaging theengaging ring 102b with firstengaging claws 62a provided on thehousing 6 of the object-capturingdevice 1. The engagingring 102b hascutout portions 102d in such a planar shape as to receive respective firstengaging claws 62a of thehousing 6. Between theengaging ring 102b and anapparatus body 10a having the recessedportion 102a formed therein, a gap G is formed to have a height large enough for receiving the firstengaging claws 62a. - In other words, when the
housing 6 is fitted into the recessedportion 102a, the firstengaging claws 62a are inserted into the recessedportion 102a throughrespective cutout portions 102d, and thehousing 6 is rotated to slide the firstengaging claws 62a into the gap G. Thereby, thehousing 6 is engaged with theengaging ring 102b. - As shown in
FIG. 3 , when the object-capturingdevice 1 is placed in the recessedportion 102a, acover 3 of the object-capturing device is removed, and a capturingdish 4 disposed in thehousing 6 is exposed, as described in detail below. - In
FIG. 3 , a throughhole 41 for injecting the reagents R (refer toFIG. 1 ) and hot water into aninternal space 66 of thehousing 6, acarrier 5 disposed on the back side of the capturingdish 4, adischarge opening 64a of thehousing 6, afilter 7 provided on the outside of thedischarge opening 64a, and asuction head 104a of the suction unit 104 (refer toFIG. 1 ) connected with thehousing 6 are shown. Theinternal space 66 corresponds to "a space" in the claims. - As shown in
FIG. 3 , the object-capturingdevice 1 is mounted on the mountingunit 102 through theengaging ring 102b. In the object-capturingdevice 1, the capturingdish 4 having the throughhole 41 is disposed above thehousing 6, and thecarrier 5 as a sample is disposed on the back side of the capturingdish 4 and in theinternal space 66 of thehousing 6. - In the object-capturing
device 1 mounted on the mountingunit 102, thehousing 6 has a protruding portion which protrudes upward from the recessedportion 102a, and the capturingdish 4 is disposed in the protruding portion. Theheater 102c may be any means that is capable of heating the recessedportion 102a (the aluminum member), which surrounds thehousing 6 of the object-capturingdevice 1 mounted on the mountingunit 102, to a predetermined temperature. Specifically, theheater 102c is preferable to be, for example, a cartridge heater. - The
functional liquid tank 105 shown inFIG. 1 is adapted to store liquid such as sterile distilled water. This liquid is poured into the housing 6 (refer toFIG. 3 ) to, for example, improve a filtration rate of the carrier 5 (refer toFIG. 3 ) of the object-capturingdevice 1, or to wash the housing, as described below. The liquid is also poured into a piping system connected to asyringe pump 106c of thedispensing unit 106 as described below. Thefunctional liquid tank 105 may store buffer solution. - The hot-
water supplying unit 103 heats and supplies, for example, the sterile distilled water supplied from thefunctional liquid tank 105. Specifically, the hot-water supplying unit 103 injects hot water into the internal space 66 (refer toFIG. 3 ) in thehousing 6 through the through hole 41 (refer toFIG. 3 ) of the capturingdish 4. For example, the hot-water supplying unit 103 is a unit (not shown) for discharging hot water heated by a cartridge heater with a peristaltic pump through a nozzle formed with, for example, a flexible tube. This nozzle having a means for moving in horizontal and vertical directions is inserted into the internal space 66 (refer toFIG. 3 ) of thehousing 6 through the through hole 41 (refer toFIG. 3 ) of the capturingdish 4 as necessary. - The
suction unit 104 shown inFIG. 1 sucks, for example, the hot water and the reagents R described below, which are injected into the internal space 66 (refer toFIG. 3 ) in thehousing 6, to discharge them through the filter 7 (refer toFIG. 3 ). For example, thissuction unit 104 includes thesuction head 104a (refer toFIG. 3 ) described above, a suction pump (not shown) connected with thesuction head 104a through predetermined piping, and a waste tank. - The
suction unit 104 to be used according to the embodiment further includes a lifting apparatus (not shown) lifting and lowering thesuction head 104a to enable thesuction head 104a (refer toFIG. 3 ) to be engaged with and disengaged from the housing 6 (refer toFIG. 3 ) mounted on the mountingunit 102. - In the reagent cartridge 2 shown in
FIG. 1 , multiple reagents R necessary to the ATP method are arranged in a block. The reagent cartridge 2 is disposed at a predetermined position in the vicinity of the mountingunit 102. In the reagent cartridge 2, each reagent R is disposed at a predetermined position, and a dispensingnozzle 106a of thedispensing unit 106 described below dispenses the reagents R in a predetermined order into thehousing 6 of the object-capturingdevice 1 and into a luminescence-test tube 107a of the luminescence-intensity measurement unit 107. In other words, the location (the coordinates) of each reagent R is stored in thecontrol unit 108 controlling thedispensing unit 106, as described below. - Examples of the reagents R necessary to the ATP method includes an ATP eliminating reagent for eliminating ATPs that exist out of cells of captured microorganisms, an ATP extracting reagent for extracting ATPs contained in the microorganisms, and an ATP luminescence reagent for producing luminescence of the ATPs extracted from the microorganisms.
- Examples of the ATP eliminating reagent include an ATP-degrading enzyme.
- Examples of the ATP extracting reagent include a benzalkonium chloride, a trichloroacetic acid, and a Tris buffer solution.
- Examples of the ATP luminescence reagent include a luciferase/luciferin reagent.
- The examples of the reagents R may include a correction reagent for the luminescence-
intensity measurement unit 107, and sterile pure water. - The dispensing
unit 106 shown inFIG. 1 dispenses the reagents R described above into thehousing 6 of the object-capturingdevice 1. The dispensingunit 106 also dispenses the reagents R and ATP extracted solution in the housing 6 (refer toFIG. 3 ) as described below into the luminescence-test tube 107a of the luminescence-intensity measurement unit 107. - The dispensing
unit 106 may include the dispensingnozzle 106a formed with a thin tube, anactuator 106b moving the dispensingnozzle 106a in the xyz axis directions, thesyringe pump 106c connected with the dispensingnozzle 106a through predetermined flexible piping, the piping (not shown) supplying, for example, sterile distilled water from thefunctional liquid tank 105 through thesyringe pump 106c to the dispensingnozzle 106a. - The luminescence-
intensity measurement unit 107 shown inFIG. 1 may be a unit which includes the luminescence-test tube 107a for receiving the ATP extracted solution dispensed from the housing 6 (refer toFIG. 3 ) as described below to produce luminescence of ATPs, and a luminescence detectingunit body 107b having, for example, a photomultiplier which detects luminescence intensity of the ATPs. - The
control unit 108 shown inFIG. 1 has overall control over themicroorganism counting apparatus 10. Thecontrol unit 108 also controls the hot-water supplying unit 103, thesuction unit 104, the dispensingunit 106, and the luminescence-intensity measurement unit 107 according to a procedure to be described below, after the object-capturing device 1 (refer toFIG. 3 ) is mounted on the mountingunit 102. Thiscontrol unit 108 includes a CPU, a ROM, a RAM, various interfaces, and circuitry. - A procedure of execution by the
control unit 108 will be described. In this description, operations of themicroorganism counting apparatus 10 and a method for counting microorganisms will be described.FIG. 4 , which is referred to in the following description, is a flowchart showing the procedure in which the microorganism counting apparatus operates based on instructions of the control unit. - In the
microorganism counting apparatus 10 shown inFIG. 1 , thecontrol unit 108 starts execution of the following procedure when a start switch (not shown) is turned on after the object-capturing device 1 (refer toFIG. 3 ) is mounted on the mountingunit 102. - As shown in
FIG. 4 , thecontrol unit 108 sends an instruction to, for example, a predetermined inverter to apply power to theheater 102c (refer toFIG. 3 ) to generate heat. On the request of thecontrol unit 108, the temperature of the carrier 5 (refer toFIG. 3 ) of the object-capturingdevice 1 is raised with theheater 102c (Step S201). Thereby, thecarrier 5 solates and falls down off the capturing dish 4 (refer toFIG. 3 ) onto the inside bottom of the housing 6 (refer toFIG. 3 ). - The
control unit 108 sends an instruction to the hot-water supplying unit 103 (refer toFIG. 1 ) to inject hot water into the housing 6 (refer toFIG. 3 ) (Step S202). Thereby, the carrier 5 (refer toFIG. 3 ) further solates, and is diluted with the hot water. - The
control unit 108 sends an instruction to the suction unit 104 (refer toFIG. 1 ) to engage thesuction head 104a (refer toFIG. 3 ) with the housing 6 (refer toFIG. 3 ), and then suck and filter the contents in the housing 6 (refer toFIG. 3 ) (Step S203). Thereby, the microorganisms captured in thecarrier 5 are separated and held by the filter 7 (refer toFIG. 3 ), and thediluted carrier 5 is filtered and discharged out of thehousing 6. - The
control unit 108 again sends an instruction to the hot-water supplying unit 103 to dispense hot water into the housing 6 (refer toFIG. 3 ) (Step S204). Then, the hot water in thehousing 6 is filtered again (Step S205). Thereby, the filtered hot water removes remaining dilutedcarrier 5 from the inside of thehousing 6, and accordingly the recovery rate of microorganisms at thefilter 7 is improved. - The
control unit 108 sends to an instruction to the dispensing unit 106 (refer toFIG. 1 ) to dispense the ATP eliminating reagent in the reagent cartridge 2 into the housing 6 (refer toFIG. 3 ) (Step S206). As a result, the ATPs that exist out of the cells of the microorganisms on thefilter 7 are eliminated. - The
control unit 108 sends an instruction to the suction unit 104 (refer toFIG. 1 ) to suck the contents of the housing 6 (refer toFIG. 3 ) and filter the sucked contents (Step S207). Thereby, the microorganisms are separated and held by the filter 7 (refer toFIG. 3 ), and the ATP eliminating reagent and the hot water are filtered and discharged out of thehousing 6. - The
control unit 108 sends an instruction to the dispensing unit 106 (refer toFIG. 1 ) to dispense the ATP luminescence reagent in the reagent cartridge 2 into the luminescence-test tube 107a (refer toFIG. 1 ) (Step S208). - The
control unit 108 sends an instruction to the luminescence-intensity measurement unit 107 (refer toFIG. 1 ) to turn on the luminescence detectingunit body 107b (refer toFIG. 1 ) (Step S209). - The
control unit 108 sends an instruction to the dispensing unit 106 (refer toFIG. 1 ) to dispense the ATP extracting reagent in the reagent cartridge 2 into the housing 6 (refer toFIG. 3 ) (Step S210). Thereby, ATPs are extracted from the microorganisms held by thefilter 7, and sample solution is prepared on thefilter 7. - After Steps S208 and S209, the luminescence detecting unit of the luminescence-
intensity measurement unit 107 performs a background measurement of the luminescence-test tube 107a with the ATP luminescence reagent. - The
control unit 108 sends an instruction to the dispensing unit 106 (refer toFIG. 1 ) to dispense an adequate amount of the sample solution (i.e., the ATP extracted solution) in thehousing 6 into the luminescence-test tube 107a where the background measurement has been performed (Step S211). Thereby, the ATP extracted solution reacts with the ATP luminescence reagent which has been dispensed in Step S208, and produces luminescence in the luminescence-test tube 107a. - The luminescence detecting
unit body 107b (refer toFIG. 1 ) of the luminescence-intensity measurement unit 107 (refer toFIG. 1 ) detects the ATP luminescence and outputs signals. Thecontrol unit 108 digitizes the outputted signals, and measures luminescence intensity based on the single-photon counting method (Step S212). Then, thecontrol unit 108 calculates the ATP amount (amol) in the ATP extracted solution dispensed into the luminescence-test tube 107a based on a prestored calibration curve indicating the relation between the ATP amount (amol) and the luminescence intensity (CPS), and thecontrol unit 108 counts the microorganisms using an ATP value, which may be converted into the equivalent number of the microorganisms in thecarrier 5. This ATP value is calculated based on the ATP amount (amol) and the amount of the ATP extracted solution of the sample solution prepared in Step S210 (Step S213). - The object-capturing
device 1 to be used according to the embodiment will be described. The up and down direction of the object-capturingdevice 1 in the following description is the same as that shown inFIG. 5. FIG. 5 is a perspective view showing the object-capturing device to be used according to the embodiment.FIG. 6A is an exploded perspective view showing the object-capturing device ofFIG. 5 viewed from obliquely above.FIG. 6B is an exploded perspective view showing the object-capturing device ofFIG. 5 viewed from obliquely below.FIG. 7 is a cross-sectional view along a line VII-VII inFIG. 5 . - The object-capturing
device 1 to be used according to the embodiment is placed in an impactor-type air sampler 50 (refer toFIG. 8 ) described below to capture microorganisms, which are air-borne objects. The object-capturingdevice 1 is mounted in themicroorganism counting apparatus 10 described above to count the captured microorganism. The object-capturingdevice 1 is turned upside down and used when microorganisms are captured, as described in detail below. - As shown in
FIG. 5 , an upper portion of the object-capturingdevice 1 is in a substantially cylindrical shape, and a lower portion of the object-capturingdevice 1 is in a substantially conical shape so that the diameter of the horizontal cross section of the object-capturingdevice 1 becomes small downward as the horizontal cross section lowers. As described in detail below, the object-capturingdevice 1 is engaged with the air sampler 50 (refer toFIG. 8 ) in the upper portion of the object-capturingdevice 1 when microorganisms are captured. The object-capturingdevice 1 is engaged with themicroorganism counting apparatus 10 in the lower portion of the object-capturingdevice 1 when the captured microorganisms are counted. - In
FIG. 5 , thecover 3, thehousing 6, secondengaging claws 31 engaging with the air sampler 50 (refer toFIG. 8 ) described below, and the firstengaging claws 62a engaging with themicroorganism counting apparatus 10 are shown. Theair sampler 50 corresponds to "a capturing apparatus which captures the object" in the claims. Each second engagingclaw 31 corresponds to "an engaging portion" in the claims. - As shown in
FIGs 6A and 6B , the object-capturingdevice 1 includes thecover 3, the capturingdish 4, thecarrier 5, thehousing 6, thefilter 7, and a filter-securingring 8, which are disposed in this order from upward to downward and fitted with each other. - The
cover 3 is attached to close an upper opening of thehousing 6 described below, and has a cylindrical shape with a bottom and an opening facing upward. On an upper circumferential edge of the outer cylindrical surface of thecover 3, the secondengaging claws 31 described above are formed to protrude radially outward, and to be disposed in a constant spacing with each other on the circumferential surface of thecover 3. The number of the secondengaging claws 31 is three, which is same as the number of engaging cutout portions 53 (refer toFIG. 8 ) described below of theair sampler 50. - On a lower circumferential edge of the outer cylindrical surface of the
cover 3, three thirdengaging claws 32 are formed to protrude radially outward, and to be disposed in a constant spacing with each other on the circumferential surface of thecover 3. The thirdengaging claws 32 are fitted in respective first L-shapedgrooves 61a described below of thehousing 6 to detachably engage thecover 3 with thehousing 6. - As shown in
FIG. 6B , an outer bottom surface of thecover 3 forms an uneven surface constituted by multiple straight ridges and straight grooves disposed alternately and in parallel with each other, the ridges of which protrude downward. When the outer bottom surface of thecover 3 contacts with an upper surface of the capturingdish 4 as described below, this uneven outer bottom surface reduces the area of contact with the capturingdish 4. For example, after the object-capturingdevice 1 is mounted on the mounting unit 102 (refer toFIG. 2 ) of themicroorganism counting apparatus 10 described above, when thecover 3 is removed from thehousing 6, this uneven surface facilitates easy detachment of thecover 3 from the capturingdish 4 which is left in thehousing 6. As described below, after microorganisms are captured using the air sampler 50 (refer toFIG. 8 ), when the object-capturingdevice 1 is carried to a microorganism counting facility (for example, a facility having the microorganism counting apparatus 10 (refer toFIG. 1 )) at low temperature as necessary, condensation may rarely occur between thecover 3 and the capturingdish 4. Even in this case, the uneven surface facilitates easy detachment of thecover 3 from the capturingdish 4. This uneven surface is not limited to the above straight ridges and straight grooves, and may be formed with multiple protrusions, or with grains such as a matte finish pattern or a texture pattern. - As shown in
FIG. 6B , on the outer bottom surface of thecover 3, aprotrusion 33 in a cylindrical shape is formed to protrude downward. Theprotrusion 33 has an outer diameter rather smaller than the inner diameter of the throughhole 41 of the capturingdish 4 to be described below. The height of theprotrusion 33 equals to that of the throughhole 41. - As shown in
FIGs 6A and 6B , the capturingdish 4 has a disk shape. The throughhole 41, which is bored through the capturingdish 4 in the vertical direction, is formed in a center portion of the capturingdish 4. - As shown in
FIG. 6A , the upper surface of the capturingdish 4 forms an even surface to be made in contact with the outer bottom surface of thecover 3 described above. - On a lower surface of the capturing
dish 4, double ring-shapedribs hole 41 in an inner portion and an outer portion, to receive the ring-shapedcarrier 5 as described below. - The outer diameter of the capturing
dish 4 ranges between the inner diameter of alower cylinder portion 62 and the inner diameter of anupper cylinder portion 61 of the housing 6 (including both end values). Preferably, the outer diameter of the capturingdish 4 is substantially equal to the inner diameter of theupper cylinder portion 61. The outer diameter of the ring-shapedrib 42b formed in the outer portion of the capturingdish 4 shown inFIG. 6B is equal to the inner diameter of thelower cylinder portion 62 described below, or less. Preferably, the outer diameter of the ring-shapedrib 42b is substantially equal to the inner diameter of thelower cylinder portion 62. - The
carrier 5 is placed in the air sampler 50 (refer toFIG. 8 ), as described below, to receive air flow when theair sampler 50 sucks the air, and to capture microorganisms carried in the air flow. - The
carrier 5 is made of a material that undergoes a phase transition from gel to sol when the temperature rises from the room temperature. The material of thecarrier 5 is preferably such a material as to undergoes a phase transition from gel to sol at 30°C or higher. More preferably, the material liquefies at a temperature between 37°C and 40°C. Most preferably, the material is a gelatin, a mixture of gelatin and glycerol, or a copolymer having a ratio of N-acryloylglycinamide to N-methacryloyl-N'-biotinyl propylene diamine of 10:1. - The
carrier 5 has a ring shape, as described above. As shown inFIG. 6B , thecarrier 5 preferably has the same shape as that of the space formed between the ring-shapedribs dish 4. - The
carrier 5 may be formed by applying the material described above to the space formed between the ring-shapedribs - As shown in
FIGs 6A and 6B , thehousing 6 has theupper cylinder portion 61 having the inner diameter substantially as large as the outer diameter of thecover 3 as described above, thelower cylinder portion 62 having the inner diameter smaller than the inner diameter of theupper cylinder portion 61, aconical portion 64 being in an inverted conical shape with an inner diameter which becomes smaller from the inner diameter of thelower cylinder portion 62, and a filterfitting portion 65 provided on the periphery of an outlet of thedischarge opening 64a formed in the lowest portion of theconical portion 64, which are disposed in this order from upward to downward to form an integral unit. - On an inner circumferential surface of the
upper cylinder portion 61, three of the first L-shapedgrooves 61a, into which the thirdengaging claws 32 of the cover are fitted, are formed at positions corresponding to the thirdengaging claws 32, as described above. - The
lower cylinder portion 62 is connected with theupper cylinder portion 61 through ashelf portion 63. - On an outer circumferential surface of the
lower cylinder portion 62, the firstengaging claws 62a are formed to be engaged withengaging ring 102b (refer toFIG. 2 ) of themicroorganism counting apparatus 10 described above. The firstengaging claws 62a protrude outward in the radial direction of thelower cylinder portion 62, and are disposed in a constant spacing with each other on the circumferential surface of thelower cylinder portion 62. The number of the firstengaging claws 62a is four. - The
conical portion 64 having the inner diameter becoming smaller downward enables the contents to easily flow down toward the lowest portion, that is, thedischarge opening 64a (refer toFIG. 6B ). - The filter
fitting portion 65 forms an integral unit with afilter housing portion 65a (refer toFIG. 6B ) forming a thin disk-shaped space, the shape of which matches that of thefilter 7 which is disposed to close the outlet of thedischarge opening 64a (refer toFIG. 6B ), and aring supporting portion 65b having a cylindrical shape and supporting the filter-securingring 8. - Second L-shaped
grooves 65c are formed on the inner circumferential surface of thering supporting portion 65b, and fourthengaging claws 82a formed on the filter-securingring 8 described below are fitted into respective second L-shapedgrooves 65c. The number of the second L-shapedgrooves 65c is four, and the second L-shapedgrooves 65c are formed to be disposed in a constant spacing with each other on the circumferential surface of thering supporting portion 65b. - The
filter 7 to be used according to the embodiment is a membrane filter. As described above, thefilter 7 closes the outlet of thedischarge opening 64a. In other words, thefilter 7 is disposed on the outside of thedischarge opening 64a. Thefilter 7 includes ahydrophilic filter 7a and ahydrophobic filter 7b, which are arranged in this order from thedischarge opening 64a. - The
hydrophilic filter 7a and thehydrophobic filter 7b may be selected from products launched in the market. Examples of thehydrophilic filter 7a include MF-Millipore (Nihon Millipore K.K.), Durapore (Nihon Millipore K.K.), and Isopore (Nihon Millipore K.K.). - Examples of the
hydrophobic filter 7b include Mitex (Nihon Millipore K.K.) and Polypropylene Prefilter (Nihon Millipore K.K.). - It should be noted that the
filter 7 used in the embodiment should have an outer diameter larger than the inner diameter of thedischarge opening 64a. - As shown in
FIGs 6A and 6B , the filter-securingring 8 fixes thefilter 7 to the housing 6 (i.e., the conical portion 64). The filter-securingring 8 has a throughhole 81 at a position where the throughhole 81 communicates with thedischarge opening 64a of theconical portion 64 through thefilter 7. - The filter-securing
ring 8 includes aring body 82 having a shape substantially same as the inner diameter of thering supporting portion 65b of thefilter fitting portion 65 described above, and aflange portion 83 formed on the lower side of thering body 82 and having a diameter larger than the outer diameter of thering body 82. - As shown in
FIG. 6A , the filter-securingring 8 further includes: afitting portion 84 which is deposited on thering body 82 so that thefitting portion 84 and thering body 82 form an integral unit, and is fitted into thefilter housing portion 65a of thehousing 6; and a ring-shapedrib 85 vertically disposed on the periphery of an opening of the throughhole 81 of thefitting portion 84. The ring-shapedrib 85 presses thefilter 7 on the periphery of the outlet of thedischarge opening 64a (refer toFIG. 6B ). - On the circumferential surface of the
ring body 82, four of the fourthengaging claws 82a are formed to protrude radially outward, and to be disposed in a constant spacing with each other on the circumferential surface of thering body 82. The fourthengaging claws 82a are formed at positions corresponding to the respective second L-shapedgrooves 65c of thering supporting portion 65b described above, and are fitted into the respective second L-shapedgrooves 65c to detachably engage the filter-securingring 8 with thehousing 6. - As shown in
FIG. 7 , the object-capturingdevice 1 as described above is formed as follows. The capturingdish 4 is mounted on theshelf portion 63 of thehousing 6; thehousing 6 is coupled to thecover 3 through the capturingdish 4 using the first L-shapedgrooves 61a and the thirdengaging claws 32; and the throughhole 41 of the capturingdish 4 is sealed by theprotrusion 33 of thecover 3. - The
housing 6 is decoupled from thecover 3 by rotating thehousing 6 relative to thecover 3 to disengage the thirdengaging claws 32 from the first L-shapedgrooves 61a. - The
filter 7 is disposed in thefilter housing portion 65a to close the outlet of thedischarge opening 64a of theconical portion 64, and thefilter fitting portion 65 is engaged with the filter-securingring 8 using the second L-shapedgrooves 65c and the fourthengaging claws 82a described above. Thereby, thedischarge opening 64a of theconical portion 64 communicates with the throughhole 81 of the filter-securingring 8 through thefilter 7. As described above, when thefilter fitting portion 65 is engaged with the filter-securingring 8, thefilter 7 is pressed by the ring-shapedrib 85 of the filter-securingring 8, and thereby thefilter 7 is disposed on the periphery of the outlet of thedischarge opening 64a. Thus, thefilter 7 is fixed firmly. - The object-capturing
device 1 as described above is used as follows. When the reagents R (refer toFIG. 1 ) described above are dispensed into thehousing 6, the throughhole 41 opens to communicate with theinternal space 66 which receives the reagents, as shown inFIG. 3 . However, before the reagents R are dispensed, theprotrusion 33 of thecover 3 seals the throughhole 41. The outlet of thedischarge opening 64a of theconical portion 64 is closed by thefilter 7 which separates microorganisms. As a result, theinternal space 66 is a space isolated from the external environment (i.e., a closed space) at least for microorganisms. Consequently, thecarrier 5 held on the capturingdish 4 is placed in this closed space. - The object-capturing
device 1 as described above other than thefilter 7 may be molded with resin, preferably polypropylene. - A method for using the object-capturing
device 1 according to the embodiment will be described. - First, a method for capturing microorganisms using the object-capturing
device 1 will be described.FIG. 8 referred to below is a perspective view showing the method for capturing microorganisms using the object-capturing device of the present invention. - As shown in
FIG. 8 , when microorganisms are captured, the object-capturingdevice 1 is used in such a way that the capturingdish 4 holding thecarrier 5 is mounted on thecover 3. In other words, the object-capturingdevice 1 shown inFIG. 7 is turned upside down and used with the capturingdish 4 left on thecover 3 and with thehousing 6 and the filter-securingring 8 removed. Thehousing 6 is removed from thecover 3 by rotating thehousing 6 relative to thecover 3 to disengage the third engaging claws 32 (refer toFIG. 6A ) from the first L-shapedgrooves 61a (refer toFIG. 6A ) as described above after thecover 3 is located in thepedestal 52 of theair sampler 50 as described below. - The object-capturing
device 1 is mounted on thepedestal 52 in a round shape in plan view, which is formed on the upper side of anair sampler body 51 of theair sampler 50. As described above, thepedestal 52 has the engagingcutout portions 53 formed to receive the secondengaging claws 31 of thecover 3, and thereby the object-capturingdevice 1 is located in a center portion of thepedestal 52. -
FIG. 8 showssuction openings 54 of theair sampler body 51, and anozzle head 55 of theair sampler 50. - According to the method for capturing microorganisms, the
housing 6 and the filter-securingring 8 which form an integral unit are removed to expose thecarrier 5 of the object-capturingdevice 1 mounted in thepedestal 52, and thenozzle head 55 is placed over thepedestal 52, as shown inFIG. 8 . The process of exposing thecarrier 5 corresponds to "the step of exposing the carrier" in the claims. - A fan (not shown) disposed in the
air sampler body 51 is activated, and the air is sucked through thesuction openings 54. Then, air flow is injected to thecarrier 5 from multiple micro nozzles (not shown) provided in thenozzle head 55. As a result, microorganisms carried in the air injected to thecarrier 5 are captured by thecarrier 5. In other words, microorganisms are captured with thecarrier 5 directed upward. - As shown in
FIG. 8 , theprotrusion 33 of thecover 3 seals the through hole 41 (refer toFIG. 7 ) of the capturingdish 4. Thus, the surface of the capturingdish 4 on the side of thecarrier 5 is flush with the bottom of theprotrusion 33. This reduces disturbance of the received air flow. Consequently, thecarrier 5 is capable of capturing microorganisms efficiently. - When the
air sampler 50 sucks a predetermined amount of the air, the process of capturing microorganisms with the object-capturingdevice 1 ends. This capturing process corresponds to "the step of capturing the object" in the claims. - When this capturing process ends, the
housing 6 and the filter-securingring 8 which form an integral unit are fitted to thecover 3 again, and the object-capturingdevice 1 returns back to the state shown inFIG. 7 . This process corresponds to "the step of placing the housing again over the capturing dish to cover the carrier" in the claims. - A method for using the object-capturing
device 1 in themicroorganism counting apparatus 10 which counts the captured microorganisms will be described. - When the capturing process described above ends, the object-capturing
device 1 as shown inFIG. 7 is carried by a user into themicroorganism counting apparatus 10, as described above (refer toFIG. 2 ). - As described above, the object-capturing
device 1 is mounted on the mountingunit 102, and thecover 3 is removed from thehousing 6 by the user (refer toFIG. 3 ). -
FIGs 9A1 to 9A4 are cross-sectional views of the object-capturing device, showing the method for using the object-capturing device in the microorganism counting apparatus.FIGs 9B1 to 9B4 are enlarged schematic diagrams showing the vicinity of the filter in the case ofFIGs 9A1 to 9A4 . -
FIGs 9B1 to 9B4 show microorganisms B and ATPs. However, the sizes of actual microorganisms are as small as in the order of micrometer, and the sizes of actual ATPs are as small as that of a molecule. Accordingly,FIGs 9B1 to 9B4 shows no relative sizes of a microorganism and an ATP. - When the temperature of the
carrier 5 is raised in Step S201 (refer toFIG. 4 ) as described above, thecarrier 5 held on the capturingdish 4 solates and falls down onto theconical portion 64 of thehousing 6 as shown inFIG. 9A1 . In this step, the microorganisms B captured with the air sampler 50 (refer toFIG. 8 ) are retained with thecarrier 5 on thefilter 7 as shown inFIG. 9B1 . - When the hot water HW is injected into the
housing 6 in Step S202 (refer toFIG. 4 ) as described above, thecarrier 5 further solates and is diluted by the hot water. Thefilter 7 includes thehydrophobic filter 7b on the lower side thereof as shown inFIG. 9A2 . Thus, the hot water HW containing the dilutedcarrier 5 is retained in thehousing 6. The microorganisms B are retained in the hot water HW containing the dilutedcarrier 5 on thefilter 7.FIG. 9A2 also shows the capturingdish 4, and the conical portion 64 (The same reference number denotes the same element throughout the following views.) - When the contents in the
housing 6 are filtered in Step S203 (refer toFIG. 4 ) as described above, the hot water HW containing the dilutedcarrier 5 in the housing 6 (refer toFIG. 9A2 ) is discharged as shown inFIG. 9A3 . In this step, the microorganisms B in the hot water HW containing the dilutedcarrier 5 are separated and held by thefilter 7 as shown inFIG. 9B3 . - As shown in
FIG. 9B3 , thefilter 7 to be used according to the embodiment has a double layer structure of thehydrophilic filter 7a and thehydrophobic filter 7b. Unlike a filter including only a hydrophilic filter, which is used in conventional ATP methods, thehydrophobic filter 7b enables liquid to be retained on the filter unless the liquid is sucked or pressure-filtered. This enables reaction with reagent, such as ATP extraction, to be performed on thefilter 7. - The ATP eliminating reagent is dispensed into the
housing 6 in Step S206 (refer toFIG. 4 ) as described above, and then the ATP extracting reagent is dispensed into thehousing 6 in Step S210 (refer toFIG. 4 ) as described above. - These processes of dispensing the reagents correspond to "the step of injecting a reagent into the housing" in the claims.
- In the
housing 6, into which the ATP extracting reagent is injected in Step S210 (refer toFIG. 4 ) as described above, ATP extracted solution EX is retained as shown inFIG. 9A4 . As shown inFIG. 9B4 , the ATP extracted solution EX contains ATPs, the amount of which corresponds to the number of the microorganisms B. - The ATP extracted solution EX shown in
FIG. 9B4 is dispensed into the luminescence-test tube 107a (refer toFIG. 1 ) in Step S211 (refer toFIG. 4 ) as described above, and then the process sequence of the method for using the object-capturingdevice 1 ends. - According to the object-capturing
device 1 and the using method thereof as described above, microorganisms are captured with thecarrier 5 directed upward, and then thecarrier 5 is directed downward to contact the microorganisms with the reagents through the throughhole 41. - Accordingly, the
carrier 5 directed upward facilitates the capturing of the microorganisms. When the reagents are contacted with the microorganisms, that is, the microorganisms are detected, thecarrier 5 is directed downward, and thereby the capturingdish 4 serves as a cover of thecarrier 5. For example, this prevents thecarrier 5 from being contaminated with, for example, dust and microbes falling from above. - According to the object-capturing
device 1 and the using method thereof, before the object-capturingdevice 1 is mounted in theair sampler 50, and during the time after the microorganisms are captured using theair sampler 50 and before the captured microorganisms are carried into themicroorganism counting apparatus 10, thecarrier 5 is placed in the closed space in thehousing 6. As a result, thecarrier 5 is prevented from being contaminated with substances which are disturbance factors for the counting of the microorganisms, unlike a conventional capturing device with an exposed carrier, such as the device disclosed in Japanese Patent Application Laid-Open No.2009-131186 - Consequently, the object-capturing
device 1 and the using method thereof enable more accurate counting of the microorganisms captured at a test site. - According to the object-capturing
device 1 and the using method thereof, when the microorganisms are contacted with the reagents R using themicroorganism counting apparatus 10, the reagents R are dispensed into thehousing 6 through the throughhole 41 of the capturingdish 4. This minimizes the communication between the inside and the outside of thehousing 6. As a result, the inside of thehousing 6 is prevented from being contaminated with substances which are disturbance factors for the counting of the microorganisms. - According to the object-capturing
device 1 and the using method thereof, thehousing 6 has the discharge opening 64a through which the contents are discharged, and thedischarge opening 64a has the filter disposed thereon to separate and hold microorganisms. Thereby, the microorganisms may be contacted with the reagents R in thehousing 6. Consequently, the object-capturingdevice 1 and the using method thereof dramatically reduce the disturbance factors for the counting of the microorganisms, unlike a conventional capturing device, such as the device disclosed in Japanese Patent Application Laid-Open No.2009-131186 - According to the object-capturing
device 1 and the using method thereof, thefilter 7 has a double layer structure of thehydrophilic filter 7a and thehydrophobic filter 7b. Thereby, reaction of reagents with recovered microorganisms may be performed on thefilter 7, unlike a filter used in the conventional ATP methods, which includes only a hydrophilic filter. - According to the object-capturing
device 1 and the using method thereof, thecover 3 has the secondengaging claws 31 formed on the opposite side of thehousing 6 to engage with the air sampler. To expose thecarrier 5, thehousing 6, which is fixed with thecover 3 to form an integral unit as shown inFIG. 5 , is grasped by hands, thecover 3 is placed into theair sampler 50 as shown inFIG. 8 , and then thehousing 6 is removed from thecover 3. In other words, when thecarrier 5 is exposed, the capturingdish 4 holding thecarrier 5 is not touched by hands. Consequently, the object-capturingdevice 1 and the using method thereof surely prevent thecarrier 5 from being contaminated with substances which are disturbance factors for the counting of the microorganism. - According to the object-capturing
device 1 and the using method thereof, after the object-capturingdevice 1 is mounted on the mountingunit 102, when thecover 3 is removed from thehousing 6 by a user, the capturingdish 4 is turned upside down relative to the state at the time when placed in the air sampler, and thereby thecarrier 5 faces toward theinternal space 66. This surely prevents the contamination of thecarrier 5. This advantage may be achieved regardless of the type or the number of thefilter 7, such as thehydrophilic filter 7a or thehydrophobic filter 7b described above. - The embodiment of the present invention has been described. The present invention is not limited to the embodiment described above, and various modifications can be made.
- According to the embodiment described above, the microorganisms captured with the object-capturing
device 1 are counted in themicroorganism counting apparatus 10. Instead of using themicroorganism counting apparatus 10, the reagents R may be manually dispensed into the housing to count the microorganisms by the ATP method. - The present invention is applicable to spore-forming bacteria such as Bacillus subtilis. In this case, examples of the reagents described above may include a vegetative cell conversion reagent, such as amino acid and sugar.
- According to the embodiment described above, the ATP method is used to count microorganisms. Instead, the microorganisms may be counted based on the fluorescence produced when substances in a living body, such as DNA, RNA, and NAD, which are extracted from the microorganisms, are irradiated with excitation light.
- In the case where the object-capturing
device 1 is used to capture and count gram negative bacilli, the counting may be made based on endotoxin contained in the cell membrane of gram negative bacilli. In other words, microbes may be counted based on luminescence intensity resulting from the limulus test on the endotoxin. - The microorganisms may be counted by recovering the microorganisms from the
filter 7 and culturing the recovered microorganisms.
Claims (1)
- A method for using an object-capturing device (1), which includes a carrier (5) capturing an object, a capturing dish (4) holding the carrier (5) on a first side of the capturing dish (4), and a housing (6) disposed relative to the capturing dish (4) to cover the carrier (5), and in which the capturing dish (4) has a through hole (41) connecting a space (66) formed between the first side and the housing (6) with the outside of the housing (6), the method comprising the steps of:exposing the carrier (5) by removing the housing (6);capturing the object onto the exposed carrier (5);placing the housing (6) again relative to the capturing dish (4) to cover the carrier (5) after the capturing of the object;solating the carrier (5) by raising the temperature of the carrier (5) and injecting hot water into the space (66) through the through hole (41);filtering the solated carrier (5); andinjecting a reagent for detecting the object, into the housing (6) through the through hole (41) formed in the capturing dish (4).
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
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JP2009295655A JP4924707B2 (en) | 2009-12-25 | 2009-12-25 | Detected object collector and method of using the same |
EP10015991.2A EP2339321B1 (en) | 2009-12-25 | 2010-12-22 | Device for capturing object |
Related Parent Applications (2)
Application Number | Title | Priority Date | Filing Date |
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EP10015991.2A Division EP2339321B1 (en) | 2009-12-25 | 2010-12-22 | Device for capturing object |
EP10015991.2A Division-Into EP2339321B1 (en) | 2009-12-25 | 2010-12-22 | Device for capturing object |
Publications (2)
Publication Number | Publication Date |
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EP2685232A1 EP2685232A1 (en) | 2014-01-15 |
EP2685232B1 true EP2685232B1 (en) | 2015-09-02 |
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Application Number | Title | Priority Date | Filing Date |
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EP10015991.2A Not-in-force EP2339321B1 (en) | 2009-12-25 | 2010-12-22 | Device for capturing object |
EP13004833.3A Not-in-force EP2685232B1 (en) | 2009-12-25 | 2010-12-22 | Method for using object-capturing device |
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Application Number | Title | Priority Date | Filing Date |
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EP10015991.2A Not-in-force EP2339321B1 (en) | 2009-12-25 | 2010-12-22 | Device for capturing object |
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EP (2) | EP2339321B1 (en) |
JP (1) | JP4924707B2 (en) |
CN (1) | CN102154096B (en) |
SG (1) | SG172584A1 (en) |
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US8628953B2 (en) * | 2007-11-29 | 2014-01-14 | Hitachi Plant Technologies, Ltd. | Capturing carrier, capturing device, analysis system using the same, and method for capturing and testing microorganisms |
JP5252176B2 (en) * | 2007-11-29 | 2013-07-31 | 株式会社日立プラントテクノロジー | Collection carrier, collection unit, collection device and collection / inspection method |
US20110018337A1 (en) * | 2008-01-11 | 2011-01-27 | General Atomics | Braking system with linear actuator |
WO2009157510A1 (en) * | 2008-06-27 | 2009-12-30 | 株式会社日立製作所 | Cartridge of microbial cell-capturing carrier, carrier treating device and method for counting microbial cells |
-
2009
- 2009-12-25 JP JP2009295655A patent/JP4924707B2/en not_active Expired - Fee Related
-
2010
- 2010-12-22 EP EP10015991.2A patent/EP2339321B1/en not_active Not-in-force
- 2010-12-22 EP EP13004833.3A patent/EP2685232B1/en not_active Not-in-force
- 2010-12-23 US US12/977,860 patent/US9476808B2/en not_active Expired - Fee Related
- 2010-12-23 SG SG2010095933A patent/SG172584A1/en unknown
- 2010-12-23 CN CN201010606111.9A patent/CN102154096B/en not_active Expired - Fee Related
Also Published As
Publication number | Publication date |
---|---|
CN102154096A (en) | 2011-08-17 |
EP2339321A2 (en) | 2011-06-29 |
US20110159536A1 (en) | 2011-06-30 |
JP2011133444A (en) | 2011-07-07 |
EP2339321B1 (en) | 2015-11-25 |
EP2685232A1 (en) | 2014-01-15 |
SG172584A1 (en) | 2011-07-28 |
US9476808B2 (en) | 2016-10-25 |
JP4924707B2 (en) | 2012-04-25 |
EP2339321A3 (en) | 2012-03-14 |
CN102154096B (en) | 2014-04-30 |
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